Due to poor prognosis and lack of effective treatment, pancreatic carcinoma (PC) is a devastating disease. With the goal of contributing to an improved detection, prevention and treatment of the disease, a comparative proteome analysis of PC and normal tissue was carried out. Paired tissue extracts from 12 patients (pancreatic adenocarcinoma and adjacent healthy tissue) were separated by two-dimensional electrophoresis. Differential protein expression was analyzed by gel comparison with the help of image analysis software. The differentially expressed spots were identified by matrix-assisted laser desorption/ionization-time of flight mass spectrometry. Seventy proteins were more strongly expressed (mostly two-fold or more) in cancerous tissue, while 41 were stronger in normal pancreas respectively. Those spots highly expressed in PC were confirmed in gels from independent individual samples. Among them were several cytoskeletal proteins, small GTP-binding proteins, and members of the S100 protein family etc. Nine proteins had been reported in previous nuclear acid-based studies. The levels of two proteins were confirmed by immunohistochemistry. One of them, fascin, was detected in 13 out of 21 carcinoma and negative in all normal pancreas samples. Moreover, fascin expression was related to the differentiation of pancreatic carcinoma.